Identification of uridylated RNA important for germline maintenance in C. elegans Fertility depends on formation and maintenance of a healthy germ line. In animals, the germ line typically derives from a very limited set of precursor cells that divide to expand the population before undergoing meiosis and gametogenesis. Maintenance of the germ line, as well as formation of high quality gametes, depends on mechanisms that ensure genome integrity as well as epigenetic processes that convey information via histone modification and small RNAs. This proposal addresses the role of poly(U) polymerase (PUP) activity in germ line maintenance and development. Poly(U) polymerases are nucleotidyl transferases that add uridine residues to the 3' end of RNA molecules. 3' uridylation is one of several post-transcriptional modifications known to modulate RNA stability. We have identified a redundant requirement for expression of two such enzymes, PUP-1 and PUP-2, in germ line survival and development in the model organism, C. elegans. PUP-1 and PUP-2 are implicated in modifying and destabilizing certain classes of small RNAs, as are their mammalian orthologs, TUT4 (ZCCHC11) and TUT7 (ZCCHC6). TUT4 and TUT7 also are known to modify mRNA, however it is not known if PUP-1 and/or PUP-2 have this activity. In this proposal, we will perform RNA sequencing experiments to identify uridylated small RNAs and mRNAs in wildtype and pup single and double mutant strains. This information will allow us to identify unique and common targets of PUP-1 and PUP-2 activity and to correlate the loss of uridylation with changes in RNA abundance. Our central hypothesis is that PUP activity modulates the abundance of small RNAs and mRNAs to ensure germ line maintenance and prevent transgenerational replicative aging of germ cells, as well as to ensure production of healthy gametes.